Study Of The Anxiolytic Effect Of Cananga Odorata Essential Oil And Its Mechanism

Ylang-ylang essential oil is extracted from the flower of tree Cananga odorata(Lamk.)Hook.f.& Thomson.It is wildly used in the perfume,skin care product and food industry for its good smell.In aromatherapy,ylang-ylang essential oil is popular for mood relaxing use.However,to our knowledge,there is a lack of scientific research to prove its effect on mood adujusting and study its function mechanism.In this study,cananga oil and ylang-ylang essential oils of different chemical constituents were collected.Several animal anxiety models were used to estimate the anxiolytic effect of ylang-ylang essential oil and the main effective compounds were identified.In addition,the change of neurotransmitters systems and cell signals in the brain that caused by ylang-ylang essential oil were studied.Main results were as follows:1.The distribution and morphological differences of genus Cananga odorata were investigated.Two groups are distinguished in cultivation: C.odorata(Lamk.)Hook.f.& Thomson.and C.odorata Baill.f.forma macrophylla.Both of them are arbors.Ylang-ylang essential oil is extracted from the former one and cananga oil is extracted from the later one.Dwarf ylang-ylang(C.odorata var.fruticosa(Craib)J.Sincl.)is a kind of shrub with fewer flowers per inflorescence and more petals.It is a variety of ylang-ylang,which can also produce essential oil but rarely be mentioned in the references and market.2.Four kinds of ylang-ylang essential oils(Mada 1,Mada 2,Com 1 and Ind 1)and one kind of cananga oil(Can 1)were collected.Gas chromatograph-mass spectrometer(GC-MS)was used to analyze their constituents.According to the results,the ylang-ylang essential oil could be classified into two types.One type includes Mada 1 and Ind 1,of which esters and alcohols were the main constituents.Benzyl benzoate and linalool were their representative compounds.Another type includes Mada 2 and Com 1,of which terpenes,like ?-caryophyllene,germacrene D and ?-farnesene,were their main constituents.The constituents of cananga oil was similar to the high terpenes type ylang-ylang essential oil but with higher contents of total terpenes and ?-caryophyllene.3.An inhalation apparatus was built and essential oil was heated to give odor exposure to the mice.Open field(OF),elevated-plus maze(EPM)and light-dark box(LDB)tests were carried out to estimate the anxiolytic effect of ylang-ylang essential oil and its impact on the locomotor of the mice.Diazepam(DZP)was used as the positive control drug.The results showed that Mada 1 played good anxiolytic effect.It significantly increased the time and entries of the mice into the open arms in the EPM test and the light area duration in the LDB test.Mada 2 and Com 1 only increased the open arm entries at a high dose in the EPM test.Cananaga oil Can 1,which has the highest content of terpenes,showed no effect on the mice in both EPM and LDB tests.These results above suggested that esters and alcohols might make important contributions to the anxiolytic effect of ylang-ylang essential oil.4.OF,EPM and LDB tests were carried out to further study the effect of ylang-ylang essential oil after acute and chronic inhalation(7-days)on both female and male mice.Mada 1(hereinafter refferred to as YYO),which showed significant anxiolytic effect in the previous study,was used in the following studies.The results showed that YYO reduced anxiety behaviors of the male but not female mice after both acute and chronic inhalation.Testosterone and 17?-estradiol were detected using ELISA methods after chronic inhalation of YYO.The results indicated that there were no differences of the two sex hormones between the control and YYO groups.YYO could also relief social anxiety.In the three-chambered social approach task,YYO increased the social contact of the test mice with the novel mice.In the pentobarbital sodium-induced sleep tests,YYO did not alter the sleep time of the mice.These results indicated that the mechanism of YYO might be different from the benzodiazepines.5.Qualitative and quantitative analyses of YYO were carried out using GC-MS.The main components were confirmed by comparsion of their retention indexes with those of standars.The actual contents of these components were calculated using an external standard method.The main constituents of the YYO were benzyl benzoate(196.6 mg/mL),linalool(139.9 mg/mL),benzyl alcohol(116.3 mg/mL),benzyl salicylate(107.1 mg/mL),geraniol(97.7 mg/mL),benzyl acetate(72.3 mg/mL)and methyl benzoate(67.3 mg/mL).OF,EPM and LDB tests were used to assess the effect of these components.The results indicated that not all of the main constituents of YYO showed anxiolytic effect;even some of them could induce anxiety at certain doses.Benzyl benzoate,benzyl alcohol and linalool were three main effective components of YYO.Benzyl benzoate significantly increased the time spent on the open arms in the EPM test and the light area duration in the LDB test.Benzyl alcohol and linalool only increased the time spent on the open arms in the EPM test while failed to affect the behaviors of the mice in the LDB test.Menthyl benzoate,geraniol and benzyl salicylate did not show anxiolytic effect and played anxiogenic effects at certain doses on mice.6.Liquid chromatogram combined with an electrochemical detector was used to detect the impact of YYO on the monoamine neurotransmitter systems in the PFC,hippocampus and striatum of the mice.The results suggested that YYO affected the 5-hydroxytryptamine(5-HT)and dopamine(DA)systems and such affection showed sexual difference.YYO elevated the 5-HT level in the hippocampus and reduced the DA content in the striatum of male mice.In contrast,YYO improved the 5-HT metabolic level in the striatum of the female mice.The impact of benzly benzoate on the neurotransmitter systems of the male mice were consistent with what the YYO performed.These results confirmed that benzyl benzoate was the major effective component of YYO.7.m-CPP,which was an agonist of 5-HT2 C receptor,was administrated to the male mice to develop an anxiety model.The behavior analysis showed that m-CPP could induce anxiety behaviors in the mice while DZP and YYO could reverse such anxiety behviours.m-CPP could reduce the 5-HT metabolism in the PFC and increased the blood plasma corticosterone level.YYO could reverse these changes.Corticosterone and 5-HT participate in the regulation of anxiety.YYO might affect the hypothalamic-pituitary-adrenal axis through regulation of the 5-HT system.Western blot was used to detect the phosphorylation levels of extracellular signal-regulated kinase 1/2(ERK1/2)and cAMP response element-binding protein(CREB)and the expression of c-Fos in the PFC and hippocampus after the EPM test.The results showed that YYO supressed the ERK1/2-CREB pathway and reduced the expression of its downstream protein c-Fos in the hippocampus.Down-regulations of ERK1/2 were also observed in the PFC.All the results above indicated that the anxiolytic effect of YYO was associated with the MAPK pathway and relevant to the serotonin system.